Cloning of Human
in a bacterial host
corresponding to the mature
& B chains of insulin are engineered into
bacterial plasmids with
an antibiotic resistance
gene and a B-gal
gene as markers.
plasmids are separately transformed
into E. coli.
Culture of the E. coli
presence of antibiotics selects those
cells that have been successfully transformed. Induction of
expression (by addition of B-galactose
sugar) also causes transcription of the insulin genes, which are then
translated by the bacterial cells.
culture is done on an industrial scale to produce thousands of
litres of cloned human insulin.
Following chemical cleavage and purification away from the plasmid
the A & B chains spontaneously assemble to
insulin. As this
process in vitro is relatively inefficient, commercial
insulin is now made by cloning the proinsulin
mRNA (without the signal peptide of pre-proinsulin) in a single host, and
specific cleavage with the proteolytic enzymes.
after © 2000
; All text
©2011 by Steven M. Carr